Research Article: Acute Inhibition of Selected Membrane-Proximal Mouse T Cell Receptor Signaling by Mitochondrial Antagonists

Date Published: November 10, 2009

Publisher: Public Library of Science

Author(s): Kwangmi Kim, Lin Wang, Inkyu Hwang, Derya Unutmaz.

Abstract: T cells absorb nanometric membrane vesicles, prepared from plasma membrane of antigen presenting cells, via dual receptor/ligand interactions of T cell receptor (TCR) with cognate peptide/major histocompatibility complex (MHC) plus lymphocyte function-associated antigen 1 (LFA-1) with intercellular adhesion molecule 1. TCR-mediated signaling for LFA-1 activation is also required for the vesicle absorption. Exploiting those findings, we had established a high throughput screening (HTS) platform and screened a library for isolation of small molecules inhibiting the vesicle absorption. Follow-up studies confirmed that treatments (1 hour) with various mitochondrial antagonists, including a class of anti-diabetic drugs (i.e., Metformin and Phenformin), resulted in ubiquitous inhibition of the vesicle absorption without compromising viability of T cells. Further studies revealed that the mitochondrial drug treatments caused impairment of specific membrane-proximal TCR signaling event(s). Thus, activation of Akt and PLC-γ1 and entry of extracellular Ca2+ following TCR stimulation were attenuated while polymerization of monomeric actins upon TCR triggering progressed normally after the treatments. Dynamic F-actin rearrangement concurring with the vesicle absorption was also found to be impaired by the drug treatments, implying that the inhibition by the drug treatments of downstream signaling events (and the vesicle absorption) could result from lack of directional relocation of signaling and cell surface molecules. We also assessed the potential application of mitochondrial antagonists as immune modulators by probing effects of the long-term drug treatments (24 hours) on viability of resting primary T cells and cell cycle progression of antigen-stimulated T cells. This study unveils a novel regulatory mechanism for T cell immunity in response to environmental factors having effects on mitochondrial function.

Partial Text: T cell activation, a series of physiological changes leading to clonal expansion and development of effector functions, commences as T cell receptor (TCR) encounters a cognate peptide in the context of major histocompatibility complex (MHC) presented by specialized immune cells called antigen presenting cells (APCs). The interaction of TCR with cognate peptide/MHC complex (pMHC) triggers a host of intracellular signaling cascades leading to cell cycle progression [1]. Despite crucial, TCR/pMHC interaction is generally insufficient for the productive T cell activation, for which accessory (costimulatory) receptor/ligand interactions, typified by CD28/B7-1 and lymphocyte function associated antigen-1 (LFA-1)/intercellular adhesion molecule-1 (ICAM-1), are required [2], [3].

While 2C T cell absorption of LdB7-1ICAM-1 pMVs proceeds via complex mechanisms, the assay for measuring extents of the pMV-absorption is simple, expeditious and statistically accurate. In addition, considering that primary resting T cells isolated from lymph nodes of TCR Tg mice along with physiological ligands expressed on biological membrane are used, results obtained using the assay may hold superior physiological relevance. Those features make the assay system ideal for systematic investigation of cellular/molecular mechanisms underlying the pMV-absorption, namely structural basis for TCR/pMHC and LFA-1/ICAM-1 interactions and ‘inside-out’ signaling for TCR-mediated LFA-1 activation, via chemical genetics approach [15].



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